Urine sample preparation is key factor in obtaining adequate and representative cells for downstream analysis. Current methodology is usually a centrifugation for separation of tumor cells from urine based on their densities. However, this method often collects all types of cells and particles, and the recovery of the cells for subsequent molecular analysis is limited. Recent advances in size-selective microfiltration technology provide an alternative approach for isolation of tumor cells from urine sample. CellSieve™ is a new microfilter membrane that has been developed by Creatv MicroTech, Inc. The CellSieve™ membrane contains a high density of pores with selective sizes, which can separate tumor cells from both blood cells and contaminated particles based on size differences. During the microfiltration process, blood cells and contaminants, which are smaller than the size of the pores, can pass through the filter membrane, whereas tumor cells, which are usually larger than the size of the pores, will be captured on the filter. The microfiltration system can produce a high recovery efficiency of tumor cells as well as a reduction in any cross-contaminations to low levels.
Analysis of tumor cells is often necessary in the diagnosis and treatment of urological cancers. In such analyses, cell morphology, protein expression and molecular alternations of the tumor cells are frequently sought to be determined. However, if urine is permitted to be collected in a clinical site and then transported to a central lab, the urine sample will often be stored at either room temperature or 4° C. for a prolonged period of hours before microfiltration and subsequent analysis can be completed. As a result, the morphology of the tumor cells in urine will frequently change due to cell lysis or apoptosis. In addition, the chemical composition of the urine sample is frequently altered upon standing as a result of environmental changes, for example, temperature change and protease digestion. Furthermore, bacterial contamination and other microorganisms may grow in urine, which may alter microfiltration and downstream analysis.
Many of the previously described preservative reagents are not compatible with the microfiltration system because they may contain organic solvents or high concentrations of acids, which may damage the microfilter membrane. For examples, methanol and glacial acetic acid can preserve some analytes in urine but they can also dissolve or damage the microfilter membrane. In addition, inclusion of methanol and high concentrations of acid comes with safety concerns to the users due to the characteristics of the hazard materials. As the results, there is a need for an effective preservative reagent that can stabilize tumor cells in urine specimen for the microfiltration.